Structure-activity relationships of N-terminal variants of peptidomimetic tissue transglutaminase inhibitors

Tissue transglutaminase (TG2) is a multifunctional protein that catalyses protein crosslinking in the extracellular matrix, and functions as an intracellular G-protein. While both activities have been associated with human diseases, its role as a G-protein has been linked to cancer stem cell survival and maintenance of a metastatic phenotype. Recently we have shown that targeted covalent inhibitors (TCIs) can react selectively with the enzyme active site of TG2, to allosterically abolish its ability to bind GTP. In the present work, we focused on the variation of the N-terminal group of these peptidomimetic inhibitors, in order to enhance efficiency, while reducing log P and the number of rotatable bonds. This approach led to the synthesis and evaluation of 41 novel inhibitors, some of which had greatly improved efficiency and affinity for TG2 (e.g. TCI 72: K_I = 1.0 μM, k_inact/K_I = 4.4 × 10⁵ M^-1 min^-1). Molecular modelling provided a hypothetical binding mode for these TCIs. The most efficient inhibitors were evaluated further and shown to have excellent isozyme selectivity, to block GTP binding, and to have improved pharmacokinetic properties, as expected. Their biological activity was also confirmed, in a cellular invasion assay, although with less potency than expected.